The timing of reproduction is a critical event in plant development and is controlled by a refined genetic network. To regulate flowering time, plants interpret a variety of signals that converge from multiple genetic pathways at the shoot apex where the reproductive transition is effected2,3. These pathways include hormone signaling, perception of day length, developmental phase and resource availability, ambient temperature, light quality, and the passage of cold, or vernalization2,3. The genetic basis for this transition has been well characterized in the dicot Arabidopsis and several grass species in the monocots2-4. The basal dicot model genus Aquilegia has clear benefits as a system for studying the genetic basis of flowering time, most notably aspects of its evolution and ecology5-6. Within flowering plants, Aquilegia is phylogenetically intermediate between Arabidopsis and the grasses, providing a critical third data point for deep phylogenetic comparisons. At the same time, the genus Aquilegia has diversified very recently, leading to several dozen phenotypically distinct species that are broadly distributed in different environments, with very low sequence diversity between species7. Thus Aquilegia represents a model adaptive radiation. The low sequence diversity coupled with large phenotypic variation not only promises to greatly facilitate the genetic identification of adaptive traits, but also to provide a rich setting for the functional study of gene networks. This project aims to better understand the connectivity of transcription factor networks by leveraging our knowledge of flowering time in current model systems. First, it aims to perform a genome-wide interrogation of the reproductive transition in Aquilegia to better understand the network controlling this adaptive trait. Genome-wide identification of chromatin state changes implicated in governing the reproductive transition will be mapped with chromatin immunoprecipitation coupled to deep sequencing (ChIP-seq) and their output will be assessed with transcriptome sequencing. Next, the genetic and biochemical functions of genes implicated in flowering time control, the Aquilegia homologs of FLOWERING LOCUS T (FT), FD, and LEAFY (LFY) will be characterized by functional assays including knockdown by viral induced gene silencing and tests of biochemical conservation in Arabidopsis, and in situ hybridization. Finally, this project will leverage very recent genome- wide transcription factor binding maps of the key integrator protein LFY, implicating thousands of loci as direct transcriptional targets. This project will winnow out the conserved in vivo binding sites that are critical for LFY function by performing genome-wide LFY ChIP-seq assays in the Aquilegia genus in addition to more distantly related species for both micro- and macro-evolutionary comparisons. In summary, the aim is to perform functional studies of evolutionary important traits in the adaptive radiation model of the Aquilegia genus, and to address basic questions of genetic connectivity of central transcription factors by leveraging current knowledge of this network and taking advantage of species across a wide range of taxa. ! PUBLIC HEALTH RELEVANCE: The connectivity of genetic regulatory networks is not well understood and there is recent indication that the direct connectivity of these networks is unexpectedly complex. Plants present highly tractable models to access gene networks in ways that are much more cumbersome and expensive in similarly complex animals. We are therefore mapping gene networks in several species to 1) understand which interactions are most important for core developmental processes and 2) better understand the basic mechanisms in evolution.